The sulfhydryl group of the active site of glyceraldehyde-3- phosphate dehydrogenase (GPDSH) can be oxidized to a sulfenic acid. The sulfenic acid form of the enzyme (GPDSOH) is an acyl phosphatase. The acyl phosphatase reaction proceeds with the formation of a sulfenyl carboxylate intermediate. GPDSOH in concert with phosphoglycerate kinase and appropriate cofactors is an ATP'ase which will catalyze an ATP-Pi exchange. The ATP'ase is inhibited by high concentrations of uncouplers of oxidative phosphorylation and the ATP-Pi exchange is inhibited by low concentrations of the same uncouplers. The acyl phosphatase reaction catalyzed by GPDSOH is inactivated by a number of nucleophiles which include antithyroid agents, N3, ascorbate, hydrazines, and primary amines. N3, hydrazines, ascorbate, and to a limited extent, benzylamine reduce GPDSOH to GPDSH. Olefins such as dihydropyran and tetrahydrophthalimide and derivatives of it inactivate the acyl phosphatase reaction catalyzed by GPDSOH by forming addition compounds with Cys-149. Dimedone also reacts with GPDSOH to form the corresponding thioether of Cys-149. The proposed research is designed to test the possibility that a sulfenyl carboxylate is the non phosphorylated high energy intermediate in oxidative phosphorylation with C14 labeled olefins. Recent experiments have shown that ATP driven reversed electron flow at Coupling Site I is inhibited by the same olefins that react with GPDSOH. In addition we have suggested a mechanism for the non-flavin amine oxidases which postulates a sulfenic acid and a sulfenamide as intermediates on the reaction pathway. We intend to test this hypothesis with the use of C14-dimedone which inactivates the non-flavin amine oxidases.